The disclosed device, method and system can be useful for purifying a number of types of products from impurities. These can include biotherapeutic proteins and bulk biochemicals. The potential market of such products exceeds $2,000,000,000 annually. The disclosed device provides a substantial reduction in manufacturing costs of such products.
Column chromatography was described by Martin and Synge in 1941. Column chromatography is the current state of the art for purifying proteins from complex mixtures. Apart from affinity chromatography (Goding J W J. (1978), there has been little innovation in the field since its introduction.
Upstream technologies for protein production has improved product yields with the development of systems that utilize plant cells, bacterial cells, insect cells, and mammalian cells having high level expression systems and growing to high cell densities. Mammalian cell culture bioreactors have greatly increased (up to 20,000 liters) and the titers are much higher today than they were in the past. Because of larger volumes, longer fermentation times, and higher cell densities, the amounts of product and associated impurities are generally greater in the bioreactor culture fluid than in the past. Thus, improvements in product separation and purification are needed. Improvements in classical chromatography devices, methods and systems are disclosed to address this need.
Column chromatography, as traditionally practiced, utilizes an insoluble resin particle (e.g., a solid matrix particle) coated with a product binding ligand that is added to a tube (column) and allowed to settle into a “packed bed”. A suitable buffer is then pumped through the bed allowing equilibration of the ligands, to allow them to bind the desired product. The solution containing the crude product, the load, is then pumped through the resin bed. The product binds to the resin ligand along with some impurities. Another buffer, the wash buffer, formulated to wash away lightly binding impurities from the product is pumped through the resin bed and discarded. This is followed by an elution buffer, a buffer that allows the product to detach from the resin ligands, is then pumped though the resin bed and the product is eluted and collected. A buffer that detaches tightly bound impurities, the strip buffer, is then pumped through the resin bed detaching any tightly binding impurities from the resin and collected as waste. Finally, a buffer that has been formulated to re-equilibrate the resin is pumped though the resin bed enabling the entire process to be repeated.
It can be understood from the preceding description that column chromatography contains inherent limitations for purification. It is labor intensive in part because it is operated one step at a time in a batchwise process. In addition, the amount of product that can be processed depends on the size of the packed bed. Also, the rate at which material is produced depends on the maximum buffer flow rate through the packed bed. The restriction of the buffer flow rate by the packed bed causes the pressure to increase as the buffer flow rate increases. The decrease in flow rate increases process time and can adversely impact process productivity.
Column chromatography also has inherent physical limitations due to the size of the chromatography column. Chromatography columns larger than one meter in diameter are very difficult to prepare. The largest columns on the market are two meters in diameter and forty centimeters high. With these dimensions, the column can accommodate 1,250 L of resin. Since the cost of protein A resin is approximately $10,000/L, the cost of a 1,250 L protein A column is exorbitant. And assuming a binding capacity of 30 g of product/L of resin (common protein A resin capacity for monoclonal antibodies), and a chromatography column with a 50 L volume, can, in a single cycle, only bind 1.5 kg of product. A 2,000 L bioreactor with an output of 10 g/L would require a column load capacity of 200 kg. This means that the 50 L column would have to run at least 13 full cycles to process a single batch of processed protein product. Such an operation can take several days and can result in a significant production bottleneck for the manufacturing process.
Therefore, it was recognized by the inventor that breakthrough innovations in the state of the art could include 1) increased scale—leading to a larger scale of operation; 2) faster processing time; 3) reduction of raw material costs; and 4) a reduction of capital equipment costs enabled by continuous process technology.
The disclosed invention advances purification methodologies of processed products and the applications for methods derived from chromatography devices significantly by enabling continuous separation, and/or purification and/or formulation while reducing manufacturing costs and process complexity for a wide range of product types including, but not limited to biotherapeutic proteins, biologics, pharmaceuticals, antibodies and isolation of blood components, protein factors and protein fractions as well as bulk biochemicals from their natural sources. The disclosed invention can be used to replace existing slow, inefficient, batch-wise processing methods with modern continuous manufacturing technology controlled by PAT (process analytical technology). Therefore, the disclosed device, methods and systems meet the need of providing easier, faster and more economical product processing technology.